Crystal (Cry) proteins are globally used in agriculture as proteinaceous insecticides. They have also been recently recognized to have great potential as anthelmintic agents in targeting parasitic roundworms (e.g. hookworms). The most extensively characterized of the anthelmintic Cry proteins is Cry5B. We report here the 2.3 Å resolution structure of the proteolytically activated form of Cry5B. This structure, which is the first for a nematicidal Cry protein, shows the familiar three-domain arrangement seen in insecticidal Cry proteins. However, domain II is unusual in that it more closely resembles a banana lectin than it does other Cry proteins. This result is consistent with the fact that the receptor for Cry5B consists of a set of invertebrate-specific glycans (attached to lipids), and also suggests that domain II is important for receptor binding. We found that not only is galactose an efficient competitor for binding between Cry5B and glycolipids, but so too is N-acetylgalactosamine (GalNAc). GalNAc is one of the core arthroseries tetrasaccharides of the Cry5B receptor, and galactose an antennary sugar that emanates from this core. These and prior data suggest that the minimal binding determinant for Cry5B consists of a core GalNAc and two antennary galactoses. Lastly, the protoxin form of Cry5B was found to bind nematode glycolipids with equal specificity as activated Cry5B, but with lower affinity. This suggests that the initial binding of Cry5B protoxin to glycolipids can be stabilized at the nematode cell surface by proteolysis. These results lay the groundwork for the design of effective Cry5B-based anthelmintics.
Crystal proteins; Bacillus thuringiensis; Nematicidal; Anthelmintic; Lectin
The MATH (meprin and TRAF-C homology) domain is a fold of seven anti-parallel β-helices involved in protein-protein interaction. Here, we report the identification and characterization of 90 MATH-domain proteins from the Brassica rapa genome. By sequence analysis together with MATH-domain proteins from other species, the B. rapa MATH-domain proteins can be grouped into 6 classes. Class-I protein has one or several MATH domains without any other recognizable domain; Class-II protein contains a MATH domain together with a conserved BTB (Broad Complex, Tramtrack, and Bric-a-Brac ) domain; Class-III protein belongs to the MATH/Filament domain family; Class-IV protein contains a MATH domain frequently combined with some other domains; Class-V protein has a relative long sequence but contains only one MATH domain; Class-VI protein is characterized by the presence of Peptidase and UBQ (Ubiquitinylation) domains together with one MATH domain. As part of our study regarding seed development of B. rapa, six genes are screened by SSH (Suppression Subtractive Hybridization) and their expression levels are analyzed in combination with seed developmental stages, and expression patterns suggested that Bra001786, Bra03578 and Bra036572 may be seed development specific genes, while Bra001787, Bra020541 and Bra040904 may be involved in seed and flower organ development. This study provides the first characterization of the MATH domain proteins in B. rapa
Brassica rapa; Phylogenetic analysis; MATH domain protein; Protein domain organization; Gene expression; Seed development.
Previous studies in narcolepsy, an autoimmune disorder affecting hypocretin (orexin) neurons and recently associated with H1N1 influenza, have demonstrated significant associations with five loci. Using a well-characterized Chinese cohort, we refined known associations in TRA@ and P2RY11-DNMT1 and identified new associations in the TCR beta (TRB@; rs9648789 max P = 3.7×10−9 OR 0.77), ZNF365 (rs10995245 max P = 1.2×10−11 OR 1.23), and IL10RB-IFNAR1 loci (rs2252931 max P = 2.2×10−9 OR 0.75). Variants in the Human Leukocyte Antigen (HLA)- DQ region were associated with age of onset (rs7744020 P = 7.9×10−9 beta −1.9 years) and varied significantly among cases with onset after the 2009 H1N1 influenza pandemic compared to previous years (rs9271117 P = 7.8×10−10 OR 0.57). These reflected an association of DQB1*03:01 with earlier onset and decreased DQB1*06:02 homozygosity following 2009. Our results illustrate how genetic association can change in the presence of new environmental challenges and suggest that the monitoring of genetic architecture over time may help reveal the appearance of novel triggers for autoimmune diseases.
Narcolepsy-hypocretin deficiency results from a highly specific autoimmune attack on hypocretin cells. Recent studies have established antigen presentation by specific class II proteins encoded by (HLA DQB1*06:02 and DQA1*01:02) to the cognate T cell receptor as the main disease pathway, with a role for H1N1 influenza in the triggering process. Here, we have used a large and well-characterized cohort of Chinese narcolepsy cases to examine genetic architecture not observed in European samples. We confirmed previously implicated susceptibility genes (T cell receptor alpha, P2RY11), and identify new loci (ZNF365, IL10RB-IFNAR1), most notably, variants at the beta chain of the T cell receptor. We found that one HLA variant, (DQB1*03:01), is associated with dramatically earlier disease onset (nearly 2 years). We also identified differences in HLA haplotype frequencies among cases with onset following the 2009 H1N1 influenza pandemic as compared to before the outbreak, with fewer HLA DQB1*06:02 homozygotes. This may be the first demonstration of such an effect, and suggests that the study of changes in GWAS signals over time could help identify environmental factors in other autoimmune diseases.
The purpose of this study was to investigate the influences of nanoscale wear particles derived from titanium/titanium alloy-based implants on integration of bone. Here we report the potential impact of titanium oxide (TiO2) nanoparticles on adhesion, migration, proliferation, and differentiation of mesenchymal stem cells (MSC) from the cellular level to the molecular level in the Wistar rat.
A series of TiO2 nanoparticles (14 nm, 108 nm, and 196 nm) were synthesized and characterized by scanning electron microscopy and transmission electron microscopy, respectively.
The TiO2 nanoparticles had negative effects on cell viability, proliferation, and the cell cycle of MSC in a dose-dependent and size-dependent manner. Confocal laser scanning microscopy was used to investigate the effects of particle internalization on adhesion, spreading, and morphology of MSC. The integrity of the cell membrane, cytoskeleton, and vinculin of MSC were negatively influenced by large TiO2 nanoparticles.
The Transwell migration assay and a wound healing model suggested that TiO2 nanoparticles had a strong adverse impact on cell migration as particle size increased (P < 0.01). Furthermore, alkaline phosphatase, gene expression of osteocalcin (OC) and osteopontin (OPN), and mineralization measurements indicate that the size of the TiO2 nanoparticles negatively affected osteogenic differentiation of MSC.
mesenchymal stem cells; titanium dioxide; nanoparticles; cytotoxicity; adhesion; migration
The genus Gossypium is a globally important crop that is used to produce textiles, oil and protein. However, gossypol, which is found in cultivated cottonseed, is toxic to humans and non-ruminant animals. Efforts have been made to breed improved cultivated cotton with lower gossypol content. The delayed gland morphogenesis trait possessed by some Australian wild cotton species may enable the widespread, direct usage of cottonseed. However, the mechanisms about the delayed gland morphogenesis are still unknown. Here, we sequenced the first Australian wild cotton species (Gossypiumaustrale) and a diploid cotton species (Gossypiumarboreum) using the Illumina Hiseq 2000 RNA-seq platform to help elucidate the mechanisms underlying gossypol synthesis and gland development. Paired-end Illumina short reads were de novo assembled into 226,184, 213,257 and 275,434 transcripts, clustering into 61,048, 47,908 and 72,985 individual clusters with N50 lengths of 1,710 bp, 1544 BP and 1,743 bp, respectively. The clustered Unigenes were searched against three public protein databases (TrEMBL, SwissProt and RefSeq) and the nucleotide and protein sequences of Gossypiumraimondii using BLASTx and BLASTn. A total of 21,987, 17,209 and 25,325 Unigenes were annotated. Of these, 18,766 (85.4%), 14,552 (84.6%) and 21,374 (84.4%) Unigenes could be assigned to GO-term classifications. We identified and analyzed 13,884 differentially expressed Unigenes by clustering and functional enrichment. Terpenoid-related biosynthesis pathways showed differentially regulated expression patterns between the two cotton species. Phylogenetic analysis of the terpene synthases family was also carried out to clarify the classifications of TPSs. RNA-seq data from two distinct cotton species provide comprehensive transcriptome annotation resources and global gene expression profiles during seed germination and gland and gossypol formation. These data may be used to further elucidate various mechanisms and help promote the usage of cottonseed.
Transcriptional effectors of white adipocyte-selective gene expression have not been described. Here we show that TLE3 is a white-selective cofactor that acts reciprocally with the brown-selective cofactor Prdm16 to specify lipid storage and thermogenic gene programs. Occupancy of TLE3 and Prdm16 on certain promoters is mutually exclusive, due to the ability of TLE3 to disrupt the physical interaction between Prdm16 and PPARγ. When expressed at elevated levels in brown fat, TLE3 counters Prdm16, suppressing brown-selective genes and inducing white-selective genes, resulting in impaired fatty acid oxidation and thermogenesis. Conversely, mice lacking TLE3 in adipose tissue show enhanced thermogenesis in inguinal white adipose depots and are protected from age-dependent deterioration of brown adipose tissue function. Our results suggest that the establishment of distinct adipocyte phenotypes with different capacities for thermogenesis and lipid storage is accomplished in part through the cell type–selective recruitment of TLE3 or Prdm16 to key adipocyte target genes.
Our recent evidence showed that Toll like receptor 9 (TLR9) signaling could enhance the growth and metastatic potential of human lung cancer cells through repressing microRNA-7 (miR-7) expression. Human antigen R (HuR) has been involved in stabilizing multiple mRNAs in cellular biology. However, whether HuR also contributed to the altered expression of miR-7 in TLR9 signaling stimulated human lung cancer cells remains to be elucidated.
The expression of HuR in human lung cancer 95D cells treated with TLR9 agonist CpG Oligonucleotides (ODNs) was detected by Real-time PCR and Western blot assay. To explore the possible role of HuR on miR-7 expression, eukaryotic expression vector encoding HuR was transiently transfected into 95D cells and then the expression of miR-7 was detected by Real-time PCR assay. Moreover, RNA interference, western blot, Real-time PCR, MTT assay, BrdU labeling, invasion assay and scratch assay were employed to examine the disrupt effect of HuR on miR-7 expression in human lung cancer cells treated with CpG ODNs. Finally, inhibitors for PI3K, Akt or Erk respectively, and western blot were performed to explore the possible signaling pathway related to HuR expression in CpG ODNs treated human lung cancer cells.
Our data showed that TLR9 agonist CpG ODNs could induce the expression of HuR in human lung cancer cells. Moreover, overexpression of HuR could reduce the expression of miR-7 in lung cancer cells. Notably, down-regulation of HuR using RNA interference restored miR-7 expression in CpG ODNs treated lung cancer cells, accompanied by enhanced growth and metastatic potential. Finally, CpG ODNs could induce HuR expression through Akt pathway.
Our findings indicated that HuR could act as regulator in regulating TLR9 signaling associated biological effect in human lung cancer cells, which might be helpful for the understanding of the potential role of HuR in tumor biology.
Toll like receptor-9; miR-7; Human antigen R; Human lung cancer cell
Vibrio parahaemolyticus AphA and OpaR are the two master quorum sensing (QS) regulators that are abundantly expressed at low cell density (LCD) and high cell density (HCD), respectively, with a feature of reciprocally gradient production of them with transition between LCD and HCD. The type VI secretion system 2 (T6SS2) gene cluster can be assigned into three putative operons, namely VPA1027-1024, VPA1043-1028, and VPA1044-1046. T6SS2 contributes to adhesion of V. parahaemolyticus to host cells.
OpaR box-like sequences were found within the upstream promoter regions of all the above three operons, while none of AphA box-like elements could be identified for them. The subsequent primer extension, LacZ fusion, electrophoretic mobility shift, and DNase I footprinting assays disclosed that OpaR bound to the promoter regions of these three operons to stimulate their transcription, while AphA negatively regulated their transcription most likely through acting on OpaR. This regulation led to a gradient increase of T6SS2 transcription with transition from LCD to HCD.
V. parahaemolyticus OpaR and AphA positively and negatively regulate T6SS2 expression, respectively, leading to a gradient elevation of T6SS2 expression with transition from LCD to HCD. T6SS2 genes are thus assigned as the QS regulon members in V. parahaemolyticus.
AIM: To investigate the feasibility and optimal approach for laparoscopic pancreas- and spleen-preserving splenic hilum lymph node dissection in advanced proximal gastric cancer.
METHODS: Between August 2009 and August 2012, 12 patients with advanced proximal gastric cancer treated in Nanfang Hospital, Southern Medical University, Guangzhou, China were enrolled and subsequently underwent laparoscopic total gastrectomy with pancreas- and spleen-preserving splenic hilum lymph node (LN) dissection. The clinicopathological characteristics, surgical outcomes, postoperative course and follow-up data of these patients were retrospectively collected and analyzed in the study.
RESULTS: Based on our anatomical understanding of peripancreatic structures, we combined the characteristics of laparoscopic surgery and developed a modified approach (combined supra- and infra-pancreatic approaches) for laparoscopic pancreas- and spleen-preserving splenic hilum LN dissection. Surgery was completed in all 12 patients laparoscopically without conversion. Only one patient experienced intraoperative bleeding when dissecting LNs along the splenic artery and was handled with laparoscopic hemostasis. The mean operating time was 268.4 min and mean number of retrieved splenic hilum LNs was 4.8. One patient had splenic hilum LN metastasis (8.3%). Neither postoperative morbidity nor mortality was observed. Peritoneal metastasis occurred in one patient and none of the other patients died or experienced recurrent disease during the follow-up period.
CONCLUSION: Laparoscopic total gastrectomy with pancreas- and spleen-preserving splenic hilum LN dissection using the modified approach for advanced proximal gastric cancer could be safely achieved.
Proximal stomach; Stomach neoplasm; Laparoscopy; Lymph node excision; Splenic hilum
Soil-transmitted helminths are parasitic nematodes that inhabit the human intestine. These parasites, which include two hookworm species, Ancylostomaduodenale and Necator americanus, the whipworm Trichuristrichiura, and the large roundworm Ascarislumbricoides, infect upwards of two billion people and are a major cause of disease burden in children and pregnant women. The challenge with treating these diseases is that poverty, safety, and inefficient public health policy have marginalized drug development and distribution to control infection in humans. Anthelmintics (anti-worm drugs) have historically been developed and tested for treatment of non-human parasitic nematodes that infect livestock and companion animals. Here we systematically compare the in vitro efficacy of all major anthelmintic classes currently used in human therapy (benzimidazoles, nicotinic acetylcholine receptor agonists, macrocyclic lactones, nitazoxanide) against species closely related to human parasitic nematodes-Ancylostoma ceylanicum, Trichurismuris, and Ascarissuum--- as well as a rodent parasitic nematode used in veterinary drug discovery, Heligmosomoidesbakeri, and the free-living nematode Caenorhabditis elegans. Extensive in vitro data is complemented with single-dose in vivo data in three rodent models of parasitic diseases. We find that the effects of the drugs in vitro and in vivo can vary greatly among these nematode species, e.g., the efficacy of albendazole is strong on A. ceylanicum but weak on H. bakeri. Nonetheless, certain commonalities of the in vitro effects of the drugs can be seen, e.g., nitazoxanide consistently shows an all-or-nothing response. Our in vitro data suggest that further optimization of the clinical efficacy of some of these anthelmintics could be achieved by altering the treatment routine and/or dosing. Most importantly, our in vitro and in vivo data indicate that the hookworm A. ceylanicum is a particularly sensitive and useful model for anthelmintic studies and should be incorporated early on in drug screens for broad-spectrum human soil-transmitted helminth therapies.
Aims: We performed this retrospective study to evaluate the value of clinicopathological factors and a novel molecular marker stathmin in predicting treatment response to neoadjuvant chemotherapy (NCT) with docetaxel-containing regimens in patients with locally advanced breast cancer. Methods: Fifty-four consecutive locally advanced patients receiving docetaxel-containing NCT between January 2006 and July 2010 in Zhejiang Cancer Hospital were included. The expression levels of estrogen receptor (ER), progesterone receptor (PgR), epidermal growth factor receptor-2 (HER-2), and p53 were detected by immunohistochemistry, while expression of stathmin mRNA was measured by Quanti-Gene assay. Results: The overall clinical objective response (cOR) rate was 75.9% (41/54) in breast. A total of 34 patients (63.0%) experienced pathological OR (pOR), with pathological complete remission (pCR) rate of 20.4% (11/54) in breast and 16.7% (9/54) in both breast and axilla. In univariate analysis, there were associations of pOR in both breast and axilla with age (p=0.054), ER status (p=0.059), subtypes (p=0.062), p53 (p=0.030), and stathmin expression (three terciles) (p=0.039). Mean expression of stathmin in pOR group was 0.410, compared with that in no response group of 0.556 (p=0.051 by Student's t-test). Similarly, a lower expression of stathmin might represent a higher pCR rate (p=0.061). Moreover, the LOWESS smoothing plot showed the same trend, that is, that tumor with a lower level of stathmin expression had a higher probability of response to docetaxel-containing NCT. After multivariate adjustment, both ER and stathmin remained significant with hazard ratio of 4.58 (95% CI: 1.11–18.94, p=0.036) and 2.94 (95% CI: 1.26–6.86, p=0.012), respectively. Conclusions: In conclusion, ER and stathmin were independent predictive factors for NCT with docetaxel-containing regimens.
Atherosclerosis is a chronic inflammatory disease and represents the major cause of cardiovascular morbidity and mortality. There is evidence that dihydrocapsaicin (DHC) can exert multiple pharmacological and physiological effects. Here, we explored the effect of DHC in atherosclerotic plaque progression in apoE−/− mice fed a high-fat/high-cholesterol diet.
Methods and Results
apoE−/− mice were randomly divided into two groups and fed a high-fat/high-cholesterol diet with or without DHC for 12 weeks. We demonstrated that cellular cholesterol content was significantly decreased while apoA1-mediated cholesterol efflux was significantly increased following treatment with DHC in THP-1 macrophage-derived foam cells. We also observed that plasma levels of TG, LDL-C, VLDL-C, IL-1β, IL-6, TNF-α and CRP were markedly decreased while plasma levels of apoA1 and HDL-C were significantly increased, and consistent with this, atherosclerotic lesion development was significantly inhibited by DHC treatment of apoE−/− mice fed a high-fat/high-cholesterol diet. Moreover, treatment with both LXRα siRNA and PPARγ siRNA made the up-regulation of DHC on ABCA1, ABCG1, ABCG5, SR-B1, NPC1, CD36, LDLR, HMGCR, apoA1 and apoE expression notably abolished while made the down-regulation of DHC on SRA1 expression markedly compensated. And treatment with PPARγ siRNA made the DHC-induced up-regulation of LXRα expression notably abolished while treatment with LXRα siRNA had no effect on DHC-induced PPARγ expression.
These observations provide direct evidence that DHC can significantly decrease atherosclerotic plaque formation involving in a PPARγ/LXRα pathway and thus DHC may represent a promising candidate for a therapeutic agent for the treatment or prevention of atherosclerosis.
Ascaris suum and Ascaris lumbricoides are two closely related geo-helminth parasites that ubiquitously infect pigs and humans, respectively. Ascaris suum infection in pigs is considered a good model for A. lumbricoides infection in humans because of a similar biology and tissue migration to the intestines. Ascaris lumbricoides infections in children are associated with malnutrition, growth and cognitive stunting, immune defects, and, in extreme cases, life-threatening blockage of the digestive tract and aberrant migration into the bile duct and peritoneum. Similar effects can be seen with A. suum infections in pigs related to poor feed efficiency and performance. New strategies to control Ascaris infections are needed largely due to reduced treatment efficacies of current anthelmintics in the field, the threat of resistance development, and the general lack of new drug development for intestinal soil-transmitted helminths for humans and animals. Here we demonstrate for the first time that A. suum expresses the receptors for Bacillus thuringiensis crystal protein and novel anthelmintic Cry5B, which has been previously shown to intoxicate hookworms and which belongs to a class of proteins considered non-toxic to vertebrates. Cry5B is able to intoxicate A. suum larvae and adults and triggers the activation of the p38 mitogen-activated protein kinase pathway similar to that observed with other nematodes. Most importantly, two moderate doses of 20 mg/kg body weight (143 nM/kg) of Cry5B resulted in a near complete cure of intestinal A. suum infections in pigs. Taken together, these results demonstrate the excellent potential of Cry5B to treat Ascaris infections in pigs and in humans and for Cry5B to work effectively in the human gastrointestinal tract.
Ascaris suum is an intestinal parasitic nematode of pigs that is very closely related to Ascaris lumbricoides, a major intestinal parasitic nematode of humans that infects more than one billion people worldwide. Because of reduced efficacy and the threat of resistance to the current small set of approved drugs to treat Ascaris infections, new treatments are needed. Here we test against A. suum infections the effectiveness of Cry5B, a nematode-killing protein made by the natural soil bacterium Bacillus thuringiensis and representing a promising new class of anthelmintics. We demonstrate for the first time that A. suum possesses the receptors to bind Cry5B and that Cry5B can kill A. suum larvae and adults in culture. Most importantly, we demonstrate that oral administration of Cry5B to pigs infected with A. suum larvae results in a near complete elimination of the infection. Given the similarities between A. suum and A. lumbricoides and the similarity between the pig and human gastrointestinal tracts, our data indicate that Cry5B has excellent potential to treat Ascaris infections in veterinary animals and in humans.
A facile one-step synthesis of CdTe quantum dots (QDs) in aqueous solution by atmospheric microwave reactor has been developed using 3-mercaptopropionic acid reduction of TeO2 directly. The obtained CdTe QDs were characterized by ultraviolet–visible spectroscopy, fluorescent spectroscopy, X-ray powder diffraction, multifunctional imaging electron spectrometer (XPS), and high-resolution transmission electron microscopy. Green- to red-emitting CdTe QDs with a maximum photoluminescence quantum yield of 56.68% were obtained.
CdTe quantum dots; 3-mercaptopropionic acid; TeO2; Microwave irradiation; Photoluminescence
Telomeres are protein–DNA structures found at the ends of linear chromosomes and are crucial for genome integrity. Telomeric DNA length is primarily maintained by the enzyme telomerase. Cells lacking telomerase will undergo senescence when telomeres become critically short. In Saccharomyces cerevisiae, a very small percentage of cells lacking telomerase can remain viable by lengthening telomeres via two distinct homologous recombination pathways. These “survivor” cells are classified as either Type I or Type II, with each class of survivor possessing distinct telomeric DNA structures and genetic requirements. To elucidate the regulatory pathways contributing to survivor generation, we knocked out the telomerase RNA gene TLC1 in 280 telomere-length-maintenance (TLM) gene mutants and examined telomere structures in post-senescent survivors. We uncovered new functional roles for 10 genes that affect the emerging ratio of Type I versus Type II survivors and 22 genes that are required for Type II survivor generation. We further verified that Pif1 helicase was required for Type I recombination and that the INO80 chromatin remodeling complex greatly affected the emerging frequency of Type I survivors. Finally, we found the Rad6-mediated ubiquitination pathway and the KEOPS complex were required for Type II recombination. Our data provide an independent line of evidence supporting the idea that these genes play important roles in telomere dynamics.
Homologous recombination is a means for an organism or a cell to repair damaged DNA in its genome. Eukaryotic chromosomes have a linear configuration with two ends that are special DNA–protein structures called telomeres. Telomeres can be recognized by the cell as DNA double-strand breaks and subjected to repair by homologous recombination. In the baker's yeast Saccharomyces cerevisiae, cells that lack the enzyme telomerase, which is the primary factor responsible for telomeric DNA elongation, are able to escape senescence and cell death when telomeres undergo repair via homologous recombination. In this study, we have performed genetic screens to identify genes that affect telomeric DNA recombination. By examining the telomere structures in 280 mutants, each of which lacks both a telomere-length-maintenance gene and telomerase RNA gene, we identified 32 genes that were not previously known to be involved in telomere recombination. These genes have functions in a variety of cellular processes, and our work provides new insights into the regulation of telomere recombination in the absence of telomerase.
The proximal chromosome 15q is prone to unequal crossover, leading to rearrangements. Although 15q11q13 duplications are common in patients with developmental delays and mental impairment, 15q aneusomies resulting in greater or equal to 4 copies of 15q11q13 are rare and no pentasomy 15q11q13 has been reported in the literature. Thus far, all reported high copy number 15q11q13 cases are from the West populations and no such study in Chinese patients have been documented. Dosage-response pattern of high copy number 15q11q13 on clinical presentations is still a subject for further study.
In this study, we characterized two Han Chinese patients with high copy number 15q11q13. Using chromosome banding, high resolution SNP-based cytogenomic array, Fluorescence in situ hybridization, and PCR-based microsatellite analysis, we identified two patients with tetrasomy 15q11q13 and pentasomy 15q11q13. Both 15q11q13 aneusomies resulted from a maternally inherited supernumerary marker chromosome 15, and each was composed of two different sized 15q11q13 segments covering the Prader-Willi/Angelman critical region: one being about 10 Mb with breakpoints at BP1 and BP5 regions on 15q11 and 15q13, respectively, and another about 8 Mb in size with breakpoints at BP1 and BP4 regions on 15q. Both patients presented with similar clinical features that included neurodevelopmental delays, mental impairment, speech and autistic behavior, and mild dysmorphism. The patient with pentasomy 15q11q13 was more severely affected than the patient with tetrasomy 15q11q13. Low birth weight was noted in patient with pentasomy 15q1q13.
To the best of our knowledge, this is the first case of pentasomy 15q11q13 and the first study of high copy number 15q11q13 in Han Chinese patients. Our findings demonstrate that patients with tetrasomy and pentasomy of chromosome 15q11q13 share similar spectrum of phenotypes reported in other high copy number 15q11q13 patients in the West, and positive correlation between 15q11q13 copy number and degree of severity of clinical phenotypes. Low birth weight observed in the pentasomy 15q11q13 patient was not reported in other patients with high copy number 15q11q13. Additional studies would be necessary to further characterize high copy number 15q11q13 aneusomies.
Chromosome 15; Cytogenomic array; Copy number; Pentasomy; Tetrasomy
Lysophosphatidic acid (LPA), a natural phospholipid, is able to modulate diverse cellular responses through LPA receptors (LPARs). Several studies have reported that LPAR2 gene expression is increased in a variety of cancer cells, suggesting that LPAR2 is involved in gastric cancer. The present study investigated the expression profiles of the LPAR and involvement of the receptor subtypes in the LPA-induced migration of gastric cancer cells using cell migration assays, RNA interference, quantitative real-time PCR and western blotting. LPAR2 was observed to be highly expressed in SGC-7901 cells, a human gastric cancer cell line, while LPAR1 and LPAR3 were not. Transient transfection with LPAR2 siRNA was observed to reduce LPAR2 mRNA in SGC-7901 cells and eliminate the LPA-induced cell migration. It was also observed that LPA-induced SGC-7901 cell migration was inhibited by the inhibitor for Gq/11 protein and p38. The results suggest that the LPAR2/Gq/11/p38 pathway regulates LPA-induced SGC-7901 cell migration. The present findings suggest that LPAR2 may be a potential target for the clinical treatment of gastric cancer.
lysophosphatidic acid receptor2 (LPAR2); Gq/11; cell migration; gastric cancer
Radiation pneumonitis (RP) is the most significant complication of acute treatment-related toxicities in lung cancer. Intensity-modulated radiotherapy (IMRT) with inverse planning enables us to achieve the desired dose distribution. However, there are many high-risk procedures associated with lung irradiation, including chemotherapy and surgery. We report a case of fatal treatment-related pneumonitis, where the patient had undergone postoperative IMRT for lung cancer. Following completion of radiotherapy, the patient developed progressive dyspnea. A chest computed tomography (CT) scan revealed the presence of diffuse reticular interstitial processes and honeycombing in both lungs. The fibrotic change in both lungs in a transverse view was compatible with low-dose irradiation of non-target organs at risk. Acute radiation pneumonitis was diagnosed. For patients receiving postoperative IMRT, low-dose irradiation volumes should be considered for lungs, as well as strict dose-volume histogram (DVH) parameters.
radiation pneumonitis; intensity-modulated radiotherapy; low-dose irradiated volume
Hookworm infections are one of the most important parasitic infections of humans worldwide, considered by some second only to malaria in associated disease burden. Single-dose mass drug administration for soil-transmitted helminths, including hookworms, relies primarily on albendazole, which has variable efficacy. New and better hookworm therapies are urgently needed. Bacillus thuringiensis crystal protein Cry5B has potential as a novel anthelmintic and has been extensively studied in the roundworm Caenorhabditis elegans. Here, we ask whether single-dose Cry5B can provide therapy against a hookworm infection and whether C. elegans mechanism-of-action studies are relevant to hookworms.
To test whether the C. elegans invertebrate-specific glycolipid receptor for Cry5B is relevant in hookworms, we fed Ancylostoma ceylanicum hookworm adults Cry5B with and without galactose, an inhibitor of Cry5B-C. elegans glycolipid interactions. As with C. elegans, galactose inhibits Cry5B toxicity in A. ceylanicum. Furthermore, p38 mitogen-activated protein kinase (MAPK), which controls one of the most important Cry5B signal transduction responses in C. elegans, is functionally operational in hookworms. A. ceylanicum hookworms treated with Cry5B up-regulate p38 MAPK and knock down of p38 MAPK activity in hookworms results in hypersensitivity of A. ceylanicum adults to Cry5B attack. Single-dose Cry5B is able to reduce by >90% A. ceylanicum hookworm burdens from infected hamsters, in the process eliminating hookworm egg shedding in feces and protecting infected hamsters from blood loss. Anthelmintic activity is increased about 3-fold, eliminating >97% of the parasites with a single 3 mg dose (∼30 mg/kg), by incorporating a simple formulation to help prevent digestion in the acidic stomach of the host mammal.
These studies advance the development of Cry5B protein as a potent, safe single-dose anthelmintic for hookworm therapy and make available the information of how Cry5B functions in C. elegans in order to study and improve Cry5B function against hookworms.
Hookworm infections are one of the great parasitic diseases of our time, infecting more than half a billion people worldwide and are a significant source of iron-deficient anemia. Although mass drug administrations to eliminate hookworms from children and pregnant women are being deployed, all the drugs for treatment we have lack full potency against the parasites and are showing signs of reduced efficacy. Crystal proteins, like Cry5B, made by Bacillus thuringiensis are as a class considered safe to vertebrates and have been shown to have efficacy against intestinal roundworms like hookworms. Here we show that the key mechanistic details of how Cry5B functions in hookworms is conserved with that of the model free-living roundworm Caenorhabditis elegans, which has implications for confirming Cry5B safety in vertebrates and for enhancing Cry5B efficacy against roundworms. Furthermore, we show that Cry5B works effectively as a single-dose drug against hookworm infections in hamsters and can be formulated to increase its efficacy, eliminating 97% of the parasites in a single dose. These results advance the development of a novel, safe single-dose therapy for hookworm infections in humans.
Nineteen carbapenem-nonsusceptible Proteus mirabilis isolates were recovered from intensive care units in the Second Affiliated Hospital of Zhejiang University during a 3-month period. The isolates showed a high level of resistance against ciprofloxacin, in addition to their resistance against the carbapenems. Pulsed-field gel electrophoresis (PFGE) analysis showed that these isolates belonged to three clonal strains. PCRs and DNA sequence analysis of the carbapenemase and other β-lactamase genes indicated that all the isolates harbored the blaKPC-2 gene. Twelve of 19 isolates harbored the plasmid-mediated quinolone resistance (PMQR) genes, both the qnrD and aac(6′)-Ib-cr genes. Eight representative isolates with high levels of quinolone resistance carried the similar mutation profiles of S83I in gyrA, E466D in gyrB, and S80I in parC. Reduced carbapenem susceptibility was transferred to Escherichia coli (EC600) in a conjugation experiment, while the quinolone resistance was not. DNA hybridization showed that qnrD was located on a plasmid of approximately 4.5 kb. In summary, large clonally related isolates of KPC-2-producing P. mirabilis emerged in a Chinese hospital, and qnrD was detected in KPC-producing P. mirabilis for the first time.
A mouse model with compromised mitochondrial fatty acid synthesis has been engineered in order to assess the role of this pathway in mitochondrial function and overall health. Reduction in the expression of mitochondrial malonyl CoA-acyl carrier protein transacylase, a key enzyme in the pathway encoded by the nuclear Mcat gene, was achieved to varying extents in all examined tissues employing tamoxifen-inducible Cre-lox technology. Although affected mice consumed more food than control animals, they failed to gain weight, were less physically active, suffered from loss of white adipose tissue, reduced muscle strength, kyphosis, alopecia, hypothermia and shortened lifespan. The Mcat-deficient phenotype is attributed primarily to reduced synthesis, in several tissues, of the octanoyl precursors required for the posttranslational lipoylation of pyruvate and α-ketoglutarate dehydrogenase complexes, resulting in diminished capacity of the citric acid cycle and disruption of energy metabolism. The presence of an alternative lipoylation pathway that utilizes exogenous free lipoate appears restricted to liver and alone is insufficient for preservation of normal energy metabolism. Thus, de novo synthesis of precursors for the protein lipoylation pathway plays a vital role in maintenance of mitochondrial function and overall vigor.
The Functional Assessment of Cancer Therapy (FACT) is one of the most commonly used self-report instruments for evaluation of health-related quality of life in oncology patients. However, cultural considerations necessitate testing of the subscales in different populations. We sought to qualitatively and quantitatively investigate the applicability and psychometric properties of the Chinese version of the FACT-Cervix (FACT-Cx) in Chinese women with cervical cancer.
Ten personal interviews were conducted in order to explore patients’ opinions about the scale and its items in depth. In addition the questionnaire was administered to 400 women with cervical cancer to test its psychometric properties. Reliability was assessed using Cronbach’s alpha coefficient and item-subscale correlation while validity was evaluated using factor analysis and known-group validity.
Some items related to sex and the ability to give birth were questioned in the personal interviews, mostly regarding their significance and acceptance in the Chinese cultural context. The Cronbach’s alphas of FACT-Cx and the subscales were greater than 0.7, except for the cervical-cancer-specific subscale which was 0.57. Factor analysis demonstrated that the FACT-G construct generally paralleled the original. There were significant differences in the FACT-Cx and some subscales between those receiving and not receiving treatment and among the patients with different performance status.
In general, psychometric properties of the Chinese version supported its use with cervical cancer patients in Mainland China. Further work is needed to improve the psychometric adequacy of the cervical-cancer-specific subscale and adjust it to cultural considerations.
Health-related quality of life; FACT-Cx; FACT-G; Chinese version; Psychometric properties; Cervical cancer
Pno1 is a protein that plays a role in proteasome and ribosome neogenesis in yeast. So far, its functions in mammalian cells have not been investigated. To understand its function in mammals, we performed in situ hybridization analysis of Pno1 expression in different development stages and generated Pno1 gene knockout (KO) and transgenic (Tg) mice lineages. The results showed early lethality of homozygous Pno1 KO lineage caused, as demonstrated in parallel by ex vivo experiments, by arrest of embryo development before compaction stage. Though, heterozygous (HET) mice with 50% of normal Pno1 mRNA concentration were fertile and showed no obvious anomalies. The lymphoid organs of HET mice were normal in size, weight and cellularity, with normal T and B cell subpopulations. TCR-triggered activation and proliferation of HET T cells were normal. Proteasome activities in HET organs were uncompromised. Tg mice with actin promoter-driven Pno1 expression were also fertile, with no apparent anomalies, although they expressed 2–5-fold higher Pno1 mRNA levels. The lymphoid organs of Tg mice were of normal size, weight and cellularity with normal T and B cell sub-populations. TCR-triggered activation and proliferation of Tg T cells were normal. Tg organs and tissues presented normal proteasome activity as did their wild type counterparts. Tagged Pno1 over-expression in L cells and density gradient fractionation established that Pno1 existed in large complexes with sedimentation rates between 20S and 26S, bigger than mature 26S proteasomes. Pno1 in fractions did not coincide with 40S or 60S ribosome subunits. Our study indicates that Pno1 is essential for cellular functions, but only a small percentage of its normal level is sufficient, and excessive amounts are neither harmful nor useful. The nature of the large complexes it associates with remains to be identified, but it is certain that they are not mature proteasomes or ribosomes.
Nuclear lamins are usually classified as A-type (lamins A and C) or B-type (lamins B1 and B2). A-type lamins have been implicated in multiple genetic diseases but are not required for cell growth or development. In contrast, B-type lamins have been considered essential in eukaryotic cells, with crucial roles in DNA replication and in the formation of the mitotic spindle. Knocking down the genes for B-type lamins (LMNB1, LMNB2) in HeLa cells has been reported to cause apoptosis. In the current study, we created conditional knockout alleles for mouse Lmnb1 and Lmnb2, with the goal of testing the hypothesis that B-type lamins are crucial for the growth and viability of mammalian cells in vivo. Using the keratin 14-Cre transgene, we bred mice lacking the expression of both Lmnb1 and Lmnb2 in skin keratinocytes (Lmnb1Δ/ΔLmnb2Δ/Δ). Lmnb1 and Lmnb2 transcripts were absent in keratinocytes of Lmnb1Δ/ΔLmnb2Δ/Δ mice, and lamin B1 and lamin B2 proteins were undetectable. But despite an absence of B-type lamins in keratinocytes, the skin and hair of Lmnb1Δ/ΔLmnb2Δ/Δ mice developed normally and were free of histological abnormalities, even in 2-year-old mice. After an intraperitoneal injection of bromodeoxyuridine (BrdU), similar numbers of BrdU-positive keratinocytes were observed in the skin of wild-type and Lmnb1Δ/ΔLmnb2Δ/Δ mice. Lmnb1Δ/ΔLmnb2Δ/Δ keratinocytes did not exhibit aneuploidy, and their growth rate was normal in culture. These studies challenge the concept that B-type lamins are essential for proliferation and vitality of eukaryotic cells.